tprovide Tchoest Perfimfeacrtiyve, Pulsmtaotenaroyf theHyapret,rtehnisgihon-tPhrogurgahm put DPNroAjectisol(aPtPioHn-PPa)nd GgenoetyicpingCharascetrevriiczeastionto memCboerers wiollf the PPH-PP. The PPH-Genetic Characterization Core facilities will be used for the following: 1) isolation and storage of DNA from selected tissues, 2) screening for variations in the DNA sequence of BMPR2 and other selected genes by Genetic Variation Screening or GVS, 3) DNA sequence analysis of BMPR2 and other selected genes, 4) genotyping for selected allelic changes in the BMPR2, CPS 1, HTR2A, NOS 1, NOS3, SLC6A4, TH and other selected genes and 5) genotyping microsatellite and SNP markers for fine mapping and genome scans. The PPH-Genetic Characterization Core requests an ABI PRISM 310 Genetic Analyzer, centrifuges, freezers, thermal cyclers and computer equipment needed to carry out, analyze and store the data from the studies listed above and to communicate with PPH-PP Projects I-III and Cores A and B. The PPH-Genetic Characterization Core will have access to a MagnaPure instrument for DNA and RNA isolation and an ABI 3700 Genetic Analyzer as needed. Laboratory personnel will carry out all the analytic steps and will develop reagents needed to scan the entire nucleotide sequences of the BMPR2 genes of 100 members of FPPH kindreds and 100 individuals affected with sporadic PPH that are collected by various projects. Thus, the PPH-Genetic Characterization Core will satisfy the tissue DNA isolation and all genotyping needs of the investigators in Projects I-III and also make available new analytic methods needed to detect and variations in BMPR2 and other genes for research problems demanding more innovative and cost effective solutions that are beyond the scope of a service laboratory. Vanderbilt University Medical Center Division of Pediatric Medical Genetics DD-2205 Medical Center North Nashville, TN 37232-2578 KEY PERSONNEL. See instructions. Use continuation pages as needed to provide the required information in the format shown below. Start with Principal Investigator. List all other key personnel in alphabetical order, last name first. Name Organization Role on Project Phillips, John A., ]II, M.D. Vanderbilt University Medical Center Core Director PHS 398 (Rev. 05/01) Page 2 7 2 Form Page 2 Number pages consecutively at the bottom throughout the application. Do not use suffices such as 3a, 3b. Core C: Genetic Characterization Core O Principal Investigator/Program Director (Last, #/st, middle): Loyd r James Ev M.D. FROM THROUGH DETAILED BUDGET FOR INITIAL BUDGET PERIOD DIRECT COSTS ONLY 7/1/2003 6/30/2004 PERSONNEL (Applican t orEanizab'on NAME John Phillips, M.D. Melissa Prince To be Announced CONSULTANT COSTS Elliot Dawson only) % DOLLAR AMOUNT RE(_UES"ED (omit cents) TYPE EFFORT INST. ROLE ON APPT ON BASE SALARY F_NGE PROJECT (monks) PROJ. SALARY RE(_UESTED BENEF_S TOTALS Principal 12 10% 166,700 41,675 6,793 48,468 Investiqator Lab Mgr 12 40% 45,125 18,050 4,621 22,671 Res Asst III 12 100% 37,500 37,500 9,563 47,063 Y 97,225 20,977 118,202 SUBTOTALS 5,000 5,000 EQUIPMENT (Itemize) ABI Prism 310 Genetic Analyzer 72,170 Eppendorf Mastercycler 5,520 Benchtop Centrifuge 7,730 Power Macintosh w/printer 5,100 Revco -70 Freezer 7t900 98r420 SUPPLIES (Itemize by category) DNA Isolation from Tissue (year 1 only) 1,800 Genotyping Modifiers Genes 7,000 GVS Analysis Supplies 64,812 DNA Sequencing Supplies 1,900 Genome Search Expense (Search will be performed in Yrs 2 & 3,costs spread over 5 yrs) 3,807 Fine Mapping (To be performed in Yrs 1,4, & 5, costs spread over 5 yrs) 3,941 General Lab Supplies 14r000 97r260 TRAVEL PATIENT CARE COSTS IOUTPATIENT ALTERATIONS AND RENOVATIONS (itemize by category) None OTHER EXPENSES (Itemize by category) Software Ujpgrades 1,000 Maintenance Agreement on ABI Prism 310 Genetic Analyzer 4,900 51900 SUBTOTAL DIRECT COSTS FOR INITIAL BUDGET PERIOD $324,782 I 0 CONSORTIUM/CONTRACTUAL IDIRECT COSTS COSTS IFACILITIEASND ADMINISTRATION COSTS 0 TOTAL DIRECT COSTS FOR INITIAL BUDGET PERIOD (Item7a,FacePage) $324,7821 SBIR/STTR Only: FIXED FEE REQUESTED PHS398(Rev.05/01) (Page) 273 Form Page 4 Numberpagesconsecutivelyat the bottom throughout the application.Do not use suffixes such as 3a, 3b.